The hormone, oxytocin, is synthesised by magnocellular neurones of the supraoptic and paraventricular nuclei and is released from the posterior pituitary gland into the circulation to trigger uterine contractions during parturition. Kisspeptin fibre density increases around the supraoptic nucleus over pregnancy and intracerebroventricular kisspeptin excites oxytocin neurones only in late pregnancy. However, the mechanism of this excitation is unknown. Here, we found that microdialysis administration of kisspeptin into the supraoptic nucleus consistently increased the action potential (spike) firing rate of oxytocin neurones in urethane‐anaesthetised late‐pregnant rats (gestation day 18–21) but not in non‐pregnant rats. Hazard analysis of action potential firing showed that kisspeptin specifically increased the probability of another action potential firing immediately after each action potential (post‐spike excitability) in late‐pregnant rats. Patch‐clamp electrophysiology in hypothalamic slices showed that bath application of kisspeptin did not affect action potential frequency or baseline membrane potential in supraoptic nucleus neurones. Moreover, kisspeptin superfusion did not affect the frequency or amplitude of excitatory postsynaptic currents or inhibitory postsynaptic currents in supraoptic nucleus neurones. Taken together, these studies suggest that kisspeptin directly activates oxytocin neurones in late pregnancy, at least in part, via increased post‐spike excitability. Key points Oxytocin secretion is triggered by action potential firing in magnocellular neurones of the hypothalamic supraoptic and paraventricular nuclei to induce uterine contractions during birth. In late pregnancy, kisspeptin expression increases in rat periventricular nucleus neurones that project to the oxytocin system. Here, we show that intra‐supraoptic nucleus administration of kisspeptin increases the action potential firing rate of oxytocin neurones in anaesthetised late‐pregnant rats, and that the increased firing rate is associated with increased oxytocin neurone excitability immediately after each action potential. By contrast, kisspeptin superfusion of hypothalamic slices did not affect the activity of supraoptic nucleus neurones or the strength of local synaptic inputs to supraoptic nucleus neurones. Hence, kisspeptin might activate oxytocin neurons in late pregnancy by transiently increasing oxytocin neuron excitability after each action potential.

Oxytocin is secreted by hypothalamic supraoptic nucleus (SON) and paraventricular nucleus (PVN) oxytocin neurons to induce uterine contractions during parturition. Increased activation of oxytocin neurons at parturition involves a network of afferent inputs that increase oxytocin neuron excitability. Kisspeptin fibre density increases around oxytocin neurons during pregnancy, and central kisspeptin administration excites oxytocin neurons only in late pregnancy. Kisspeptin signals via extracellular regulated kinase 1/2 (ERK1/2) and p38. Therefore, to determine whether kisspeptin excites oxytocin neurons via ERK1/2-p38 signalling in late-pregnant rats, we performed immunohistochemistry for phosphorylated ERK1/2 (pERK1/2) and phosphorylated p38 (p-p38) in oxytocin neurons of non-pregnant and late-pregnant rats. Intracerebroventricular (ICV) kisspeptin administration (2 µg) did not affect pERK1/2 or p-p38 expression in SON and PVN oxytocin neurons of non-pregnant or late-pregnant rats. Furthermore, ICV kisspeptin did not affect pERK1/2 or p-p38 expression in brain areas with major projections to the SON and PVN: the nucleus tractus solitarius, rostral ventrolateral medulla, locus coeruleus, dorsal raphe nucleus, organum vasculosum of the lamina terminalis, median preoptic nucleus, subfornical organ, anteroventral periventricular nucleus, periventricular nucleus and arcuate nucleus. Hence, kisspeptin-induced excitation of oxytocin neurons in late pregnancy does not appear to involve ERK1/2 or p38 activation in oxytocin neurons or their afferent inputs.

Oxytocin is secreted by hypothalamic supraoptic nucleus (SON) and paraventricular nucleus (PVN) oxytocin neurons to induce uterine contractions during parturition. Increased activation of oxytocin neurons at parturition involves a network of afferent inputs that increase oxytocin neuron excitability. Kisspeptin fibre density increases around oxytocin neurons during pregnancy, and central kisspeptin administration excites oxytocin neurons only in late pregnancy. Kisspeptin signals via extracellular regulated kinase 1/2 (ERK1/2) and p38. Therefore, to determine whether kisspeptin excites oxytocin neurons via ERK1/2-p38 signalling in late-pregnant rats, we performed immunohistochemistry for phosphorylated ERK1/2 (pERK1/2) and phosphorylated p38 (p-p38) in oxytocin neurons of non-pregnant and late-pregnant rats. Intracerebroventricular (ICV) kisspeptin administration (2 µg) did not affect pERK1/2 or p-p38 expression in SON and PVN oxytocin neurons of non-pregnant or late-pregnant rats. Furthermore, ICV kisspeptin did not affect pERK1/2 or p-p38 expression in brain areas with major projections to the SON and PVN: the nucleus tractus solitarius, rostral ventrolateral medulla, locus coeruleus, dorsal raphe nucleus, organum vasculosum of the lamina terminalis, median preoptic nucleus, subfornical organ, anteroventral periventricular nucleus, periventricular nucleus and arcuate nucleus. Hence, kisspeptin-induced excitation of oxytocin neurons in late pregnancy does not appear to involve ERK1/2 or p38 activation in oxytocin neurons or their afferent inputs.

Oxytocin is secreted from the posterior pituitary gland by oxytocin neurons to trigger uterine contractions during birth. We have recently shown that the oxytocin neuron firing rate increases in response to intracerebroventricular (ICV) kisspeptin only in late‐pregnant rats. Furthermore, the kisspeptin projection from the periventricular nucleus of the hypothalamus (PeN) to the perinuclear zone (PNZ) surrounding the supraoptic nucleus (SON) increases in late pregnancy. This study aimed to determine the site and mechanism of central kisspeptin activation of oxytocin neuron activity in late pregnant rats. First, in vivo extracellular single unit recordings were made from SON neurons in urethane‐anaesthetised non‐pregnant and late‐pregnant rats during microdialysis application of kisspeptin (100 μM in the dialysate) into the SON. Intra‐SON kisspeptin consistently increased the firing rate of oxytocin neurons in late‐pregnant rats but not in non‐pregnant rats. Two‐way repeated measures (RM) ANOVA revealed a significant interaction between REPRODUCTIVE STATUS and TIME (F(6,88) = 10.31, p < 0.001); post hoc all pairwise Holm‐Sidak tests revealed a significant increase in firing rate in late‐pregnant rats at 30 (p < 0.05), 40, 50 and 60 minutes (all p < 0.001 versus pre‐kisspeptin), and a significantly higher firing rate than in time‐matched recordings from non‐pregnant rats at 50 and 60 minutes (p < 0.05). Then, to examine whether excitatory effects of central kisspeptin emerges directly within the SON and paraventricular nucleus (PVN) containing oxytocin neurons, we performed single label diaminobenzidine (DAB) immunostaining against phosphorylated extracellular regulated kinase (pERK), a second messenger activated by kisspeptin. ICV cannulae were implanted on day 13 or 14‐pregnant and non‐pregnant rats and one week later, day 21 pregnant and non‐pregnant rats were perfused 15 min after ICV administration of kisspeptin (2 μg/2 μl) or vehicle under pentobarbital anaesthesia. ICV kispeptin injection did not alter pERK expression in the SON or PVN. Two‐way RM ANOVA revealed a significant effect of REPRODUCTIVE STATUS (F(1,28) = 4.78, p = 0.03) but no significant effect of TREATMENT (kisspeptin/vehicle) (F(1,28) = 0.04, p = 0.82) and no REPRODUCTIVE STATUS X TREATMENT interaction (F(1,28) = 0.81, p = 0.37 ) in the SON. There was no significant effect of REPRODUCTIVE STATUS (F(1,27) = 0.93, p = 0.34, two‐way RM ANOVA) or TREATMENT (F(1,27) = 0.93, p = 0.34) and no REPRODUCTIVE STATUS X TREATMENT interaction (F(1,27) = 0.29, p = 0.59) in the PVN. Similarly, there was no significant effect of REPRODUCTIVE STATUS (F(1,27) = 2.28, p = 0.14, two‐way RM ANOVA) or TREATMENT (F(1,27) = 0.28, p = 0.59) and no REPRODUCTIVE STATUS X TREATMENT interaction (F(1,27) = 0.07, p = 0.78) in the magnocellular division of PVN. Hence, it appears that kisspeptin excites oxytocin neurons at the end of pregnancy by a presynaptic action at local nerve terminals to stimulate peripheral oxytocin release and so might contribute to oxytocin neuron act...